Gene markers for chronic mucosal injury

ABSTRACT

The invention provides gene markers for chronic mucosal injury and ulcerative colitis. Expression products of the REG gene family can be used to detect the presence of chronic mucosal injury in a body sample of a human. The expression products of a gene represented by a Hs. 111244 polynucleotide can be used to detect ulcerative colitis in a body sample of a human. Further, these markers can be used to differentiate humans with chronic mucosal injury from humans with common acute inflammatory colon disorder, common non-inflammatory benign colon disorder, and healthy colons. The degree of injury to the colon from chronic mucosal injury can be determined and the efficacy of therapy for chronic mucosal injury can be monitored. A method of screening compounds for anti-chronic mucosal injury and anti-ulcerative activity is also provided by these gene markers.

BACKGROUND OF THE INVENTION

[0001] Clinical assessment of disease activity in ulcerative colitis orCrohn's disease is very difficult. Patient symptoms do not necessarilycorrelate with the inflammatory (disease) activity in the smallintestine and colon, leading to educated guesses being used to directanti-inflammatory therapy. Similar difficulty exists in measuring ortesting the efficacy of new therapeutic compounds. Currently the goldstandard in diagnosing ulcerative colitis or Crohn's disease is the useof fiberoptic endoscopy coupled with multiple biopsies and pathologicanalysis. This very expensive approach requires a skilled specialist andhas associated risks, such as risk of sedation, bleeding, and colonperforation. The patient is also subjected to discomfort from theprocedure and preparation.

[0002] A less invasive and less risky assessment of mucosal diseaseactivity is needed to accurately guide treatment and to provide anobjective measure of mucosal injury, both for patients and for use inclinical studies. There is also a need for a simple test to aid in thedifferentiation of chronic inflammatory disease (UC or CD) from commonacute inflammatory disorders or common non-inflammatory benigndisorders. There is a further need for a simple method for thedifferentiation of ulcerative colitis and Crohn's disease because thesurgical and medical management for these two diseases is profoundlydifferent.

SUMMARY OF THE INVENTION

[0003] It is an object of the invention to provide a method foridentifying chronic mucosal injury in a human.

[0004] It is another object of the invention to provide a method ofdetermining the degree of injury to the small intestine or colon of ahuman with chronic mucosal injury.

[0005] It is yet another object of the invention to provide a method formonitoring the efficacy of therapy for chronic mucosal injury.

[0006] It is a further object of the invention to provide a method ofscreening compounds for anti-chronic mucosal injury or anti-ulcerativecolitis activity.

[0007] These and other objects of the invention are provided by one ormore of the embodiments described below.

[0008] One embodiment of the invention provides a method of diagnosingchronic inflammatory bowel disease. At least one gene expression productof the regenerating (EG) gene family is detected in a body sample of ahuman who is suspected of having chronic inflammatory bowel disease. Thehuman is identified as having chronic inflammatory bowel disease if thegene expression product is detected.

[0009] A further embodiment of the invention provides a method to aid inthe differentiation of chronic mucosal injury from common acuteinflammatory colon disorder and common non-inflammatory benign colondisorder in a human with symptoms of bowel disease. The amount of atleast one gene expression product of the REG gene family in a bodysample of a first human who is suspected of having bowel disease, iscompared with the amount of the gene expression product in a body sampleof a second human who is healthy. The first human is identified ashaving chronic mucosal injury if the body sample of the first humancontains more of the gene expression product than the body sample of thesecond human.

[0010] Another embodiment of the invention provides a method todetermine degree of injury to small intestine or colon tissue of a humanwith chronic mucosal injury. A quantity of a gene expression product ofthe REG gene family in a body sample of a human having chronic mucosalinjury is determined. The amount is correlated with the degree of injuryto the small intestine or colon.

[0011] Still another embodiment of the invention provides a method ofmonitoring the efficacy of therapy for chronic mucosal injury in a humanbody sample. At least one gene expression product of the REG gene familyis quantitated in a body sample of a human who has been subjected totherapy for chronic mucosal injury. The quantity of the expressionproduct in the sample is compared to the quantity of the gene expressionproduct in a matched body sample of the human at an earlier time. Areduction in the quantity of the gene expression product after therapyis an index of efficacy of the therapy.

[0012] Another embodiment of the invention provides a method ofscreening compounds for anti-chronic mucosal injury activity. A coloniccell expressing a gene which is a member of the REG gene family iscontacted with a test compound. The expression of the REG gene isquantitated. A test compound which decreases expression of the gene isidentified as a potential compound for treating chronic mucosal injury.

[0013] A further embodiment of the invention provides a method ofdiagnosing ulcerative colitis. An mRNA which is expressed by a generepresented by a Hs.111244 polynucleotide is detected in a body sampleof a first human who is suspected of having ulcerative colitis. Thehuman is identified as having ulcerative colitis if the mRNA isdetected.

[0014] Still another embodiment of the invention provides a method toaid in the differentiation of ulcerative colitis from common acuteinflammatory colon disorder, common non-inflammatory benign colondisorder, and Crohn's disease in a human with symptoms of bowel disease.The amount of mRNA which is expressed by a gene represented by aHs.111244 polynucleotide in a body sample of a first human suspected ofhaving bowel disease is compared with the amount of the mRNA in acomparable body sample of a second human who is healthy. A body sampleof the first human which contains more of the mRNA than the body sampleof the second human identifies the first human as having ulcerativecolitis.

[0015] Another embodiment of the invention provides a method todetermine the degree of injury to small intestine or colon tissue of ahuman with ulcerative colitis. A quantity of an mRNA which is expressedby a gene represented by a Hs.111244 polynucleotide in a body sample ofa first human having ulcerative colitis is determined. The quantity ofthe mRNA is correlated with the degree of injury to the small intestineor colon.

[0016] Even another embodiment of the invention provides a method ofmonitoring the efficacy of therapy for ulcerative colitis in a humanbody sample. An mRNA which is expressed by a gene represented by aHs.111244 polynucleotide is quantitated in a body sample of a human whohas been subjected to therapy for ulcerative colitis. The quantity ofthe mRNA in the sample is compared to the quantity of the mRNA in amatched body sample of the human at an earlier time. A reduction in thequantity of the mRNA after therapy is an index of efficacy of thetherapy.

[0017] Still another embodiment of the invention provides a method ofscreening compounds for anti-ulcerative colitis activity. A colonic cellexpressing an mRNA which is expressed by a gene represented by aHs.111244 polynucleotide is contacted with a test compound. Theexpression of the mRNA by the cell is quantitated. A test compound whichdecreases expression of the mRNA is identified as a potential compoundfor treating ulcerative colitis.

BRIEF DESCRIPTION OF THE DRAWINGS

[0018]FIG. 1 demonstrates the hybridization signal of spiked controls.The fluorescence intensity for different levels of gene expression wasstandardized by spiking a known amount of control genes.

[0019]FIG. 2 demonstrates the expression of PSP, PAP, and REGH ininflammatory bowel disease.

[0020]FIG. 3 demonstrates the use of reverse transcriptase PCR withprimers specific for PAP, PSP, or REGH on mRNA isolated from a healthyhuman and a human with ulcerative colitis.

[0021]FIG. 4 demonstrates the purification of recombinant PSP and REGHproteins.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0022] The inventor has discovered that chronic mucosal injury can bediagnosed by detecting expression levels of the REG gene family and agene represented by a Hs.111244 polynucleotide in a human body sample.The members of the REG gene family have been found to be stronglyexpressed in regions of the colon involved with chronic mucosal injuryand in the small intestine and colon of humans with Crohn's disease.Additionally, a gene represented by the Hs.111244 polynucleotide isstrongly expressed in the colon of humans with ulcerative colitis. Suchstrong expression is both surprising and useful because the REG genefamily and the gene represented by the Hs.111244 polynucleotide are notexpressed, or are expressed at low levels, in healthy small intestineand colon tissue. Further, the expression products of the REG genefamily and the gene represented by the Hs.111244 polynucleotide can bedetected in the serum of humans with chronic mucosal injury andulcerative colitis, respectively.

[0023] Chronic mucosal injury can be caused by inflammatory boweldiseases such as ulcerative colitis and Crohn's disease. Crohn's diseaseaffects both the small intestine and the colon. Chronic mucosal injurycan further be caused by immunodeficiencies, such as chronicgranulamatous disease and transplantation rejection, and infections,such as mycobacteria.

[0024] At the present time, the human regenerating (REG) gene family isknown to contain four genes: pancreatic stone protein (PSP) (the proteinis also known as thread protein, lithostathine, and Reg) as shown in SEQID NO:1, pancreatitis-associated protein (PAP) as shown in SEQ ID NO:2,human pancreatic beta cell growth factor, also known as (INGAP), asshown in SEQ ID NO:3, and regenerating gene homologue (REGH) as shown inSEQ ID NO:4. In normal circumstances, these genes are regionallyexpressed in low amounts in the small bowel and pancreatic epithelium.Healthy colonic mucosa and small intestine has little or no expressionof the REG gene family. Ulcerative colitis, Crohn's disease, or otherchronic mucosal injury leads to high levels of REG gene expression inthe colonic mucosal or small intestine or both. This expression of themembers of the REG gene family correlates with the degree ofhistopathological injury, and is not seen in the setting of acuteself-limited colonic inflammation or common non-inflammatory benigncolon disorders.

[0025] The nucleic acid sequence of the expressed sequence tag (EST)Hs.111244 is shown in SEQ ID NO:5. The nucleic acid sequence ofHs.111244 has been newly determined and represents a more completesequence of Hs.111244 than has been previously published. This nucleicacid sequence is referred to herein as the Hs.111244 polynucleotide. Thegene represented by the expressed sequence tag (EST) Hs.111244, is notexpressed or is expressed at low levels in the healthy colon mucosa,acute self-limited colonic inflammation or common non-inflammatorybenign colon disorders, or mucosa affected by Crohn's disease. However,the gene represented by the Hs.111244 polynucleotide is expressed athigh levels in colonic mucosa affected by ulcerative colitis.

[0026] In humans who have been diagnosed with a bowel disease, detectionof levels of at least one gene expression product of the REG gene familyin a body sample can be used to diagnose or prognose chronic mucosalinjury or to monitor treatment of chronic mucosal injury. The bodysample is obtained from a human and can be, for example, a tumor, asolid tissue such as colon or small intestine tissue, or a fluid samplesuch as blood, serum, or plasma. The human from whom the body sample isobtained can be apparently healthy or can already be identified ashaving chronic mucosal injury. A comparable body sample is a body sampleobtained from a second human which is the same type of body sample asobtained from a first human. A matched body sample is a body sampleobtained from the first human at an earlier time which is the same typeof body sample from the first human obtained at a later time.

[0027] Expression products of the REG gene family can be detected in abody sample. Detection of the expression products in a human's bodysample indicates the presence of chronic mucosal injury in the human. Inone embodiment, the body sample is assayed for the presence of at leastone REG gene family protein. A REG gene family protein or polypeptide,can be detected using, for example, anti-REG gene family-specificantibodies. The antibodies can be labeled, for example, with aradioactive, fluorescent, biotinylated, or enzymatic tag and detecteddirectly, or can be detected using indirect immunochemical methods,using a labeled secondary antibody. The presence of REG gene familyprotein or polypeptides can be assayed, for example, in tissue sectionsby immunocytochemistry, or in lysates, using Western blotting, as isknown in the art. Further, REG gene family proteins or polypeptides canbe assayed by immunoprecipitation assay, enzyme-linked immunoabsorbantassay, quantitative antigen capture-based immunoassay, andradioimmunoassay.

[0028] The level of at least one REG gene family protein or polypeptidein a body sample of a human suspected of having a chronic mucosal injurycan be compared with the level of the protein or polypeptide in ahealthy body sample. The level of a REG gene family protein orpolypeptide in a body sample of a human suspected of having chronicmucosal injury can be determined using antibodies specific for the REGgene family protein or polypeptide. The level of the REG gene familyprotein or polypeptide in a healthy body sample can also be determined.The two levels are compared to each other and a higher level of the REGgene family protein or polypeptide in the suspect human's body sample ascompared to the healthy human's body sample indicates the presence ofchronic mucosal injury in the suspect human. Preferably, the increasedlevel of the REG gene family protein in the suspect sample is at least25%, 50%, 100%, 150%, 200% or 250% higher than in the healthy bodysample.

[0029] Alternatively, the presence of mRNA expressed from at least onemember of the REG gene family or mRNA expressed from the generepresented by the Hs.111244 polynucleotide can be detected in a bodysample. Detection of mRNA expressed from at least one member of the REGgene family in a body sample of a human indicates the presence ofchronic mucosal injury in the human. Detection of mRNA which isexpressed by a Hs.111244 polynucleotide in a body sample of a humansuspected of having bowel disease indicates the presence of ulcerativecolitis in the human.

[0030] mRNA expressed from the REG gene family or the gene representedby a Hs.111244 polynucleotide can be detected by any means know in theart. For example, one can use in situ hybridization in tissue sectionsor Northern blots containing poly A⁺ mRNA. Other techniques such as highdensity DNA array hybridization, ribonuclease protection assay, andserial analysis of gene expression can also be used. REG gene family- orHs.111244-specific oligonucleotide probes can be generated using thepolynucleotide sequences of the REG gene family or of the generepresented by a Hs.111244 polynucleotide. The probes are preferably atleast 12, 14, 16, 18, 20, 22, 24, or 25 nucleotides in length and can beless than 2, 1, 0.5, 0.1, or 0.05 kb in length. The probes, for example,can be synthesized chemically, generated from longer polynucleotidesusing restriction enzymes, or amplified enzymatically. The probes can belabeled, for example, with a radioactive, biotinylated, or fluorescenttag. A mixture of probes can also be used. Such mixture can contain aplurality of probes which are specific to different REG family genes orspecific for the gene represented by the Hs.111244 polynucleotide.Alternatively, each of a plurality of probes can be used separately.

[0031] One of skill in the art can readily determine differences in theamount of REG gene family mRNA or a gene represented by a Hs.111244polynucleotide mRNA transcripts between two body samples, for example,using Northern blots and nucleotide probes. The level of mRNA expressedfrom of at least one member of the REG gene family or the generepresented by the Hs.111244 polynucleotide in a body sample of a humansuspected of having chronic mucosal injury, can be compared with themRNA expression from at least one member of the REG gene family or thegene represented by Hs.111244 polynucleotide in a healthy body sample.This can be done, for example, using in situ hybridization in tissuesection or in Northern blots containing poly A⁺ mRNA. A higher level ofmRNA expressed from a gene represented by a Hs.111244 polynucleotide inthe suspect body sample as compared to the healthy body sample isindicative of ulcerative colitis in the suspect human who has providedthe body sample. A higher level of mRNA expressed from a REG family genein the suspect body sample as compared to the healthy body sample isindicative of chronic mucosal injury in the suspect human who hasprovided the body sample. Preferably, the increased level of mRNAexpressed from a member of the REG gene family or the gene representedby the Hs.111244 polynucleotide in the suspect body sample is at least25%, 50%, 100%, 150%, 200%, or 250% higher than in the healthy bodysample.

[0032] If desired, the level of a particular mRNA, polypeptide, orprotein expressed from a REG gene family member or mRNA expressed from agene represented by a Hs.111244 polynucleotide in a body sample can bequantitated. Quantitation can be accomplished, for example, by comparingthe level of expression product detected in the body sample with thelevel of expression product present in a standard curve. A comparisoncan be made visually or using a technique such as densitometry, with orwithout computerized assistance.

[0033] In a preferred embodiment, chronic mucosal injury can bedifferentiated from common acute inflammatory colon disorder and commonnon-inflammatory benign colon disorder in a human with symptoms of boweldisease. The amount of at least one gene expression product such as mRNAor protein of the REG gene family in the suspect body sample is comparedto the amount of the same gene expression product in a body sample of ahuman which is healthy. The gene expression products in the two samplescan be compared by any means known in the art. A body sample from ahuman suspected of having bowel disease which contains more of the geneexpression product than the body sample of the healthy human identifiesthe suspect human as having chronic mucosal injury. Preferably theamount of the gene expression product in the body sample of the humanwith chronic mucosal injury is increased by at least 25%, 50%, 75%,100%, 200%, or 250%.

[0034] Further, ulcerative colitis can be differentiated from commonacute inflammatory colon disorder, common non-inflammatory benign colondisorder, and Crohn's disease in a human with Hs.111244 polynucleotidein a first body sample of a human suspected of having bowel disease iscompared with the amount of the mRNA in a body sample of a second humanwhich is healthy. The amount of mRNA in the two samples can be comparedby any means known in the art. A body sample from a human suspected ofhaving bowel disease which contains more of the mRNA than the bodysample of the healthy human identifies the suspect human as havingulcerative colitis. Preferably the amount of mRNA in the body sample ofthe human with ulcerative colitis is increased by at least 25%, 50%,75%, 100%, 200%, or 250%.

[0035] The degree of injury to the small intestine or colon tissue of ahuman with chronic mucosal injury can be determined by measuring thequantity of a gene expression product, such as mRNA or protein, of theREG gene family in a body sample of the human. The quantity of the geneexpression product is correlated with the degree of injury to the smallintestine or colon.

[0036] Further the degree of injury to the colon tissue of a human withulcerative colitis can be determined by measuring the quantity of a mRNAwhich is expressed by a Hs.111244 polynucleotide or the gene representedby it in a body sample of the human. The quantity of the mRNA iscorrelated with the degree of injury to the colon.

[0037] The efficacy of therapy for chronic mucosal injury can bemonitored in a body sample of a human with chronic mucosal injury. Atleast one gene expression product of the REG gene family can bequantitated in a body sample of a human which has been subjected totherapy for chronic mucosal injury. The quantity of the gene expressionproduct in a matched body sample is compared to the quantity of the geneexpression product in the body sample at an earlier time. A reduction ofin the quantity of the gene expression product after therapy is an indexof efficacy of the therapy. Preferably, the amount of the geneexpression product is decreased by at least 10%, 25%, 50%, 75% or 100%.

[0038] The efficacy of therapy for ulcerative colitis in a human bodysample can also be monitored. An mRNA which is expressed by a Hs.111244polynucleotide or the gene represented by it is quantitated in a bodysample of a human which has been subjected to therapy for ulcerativecolitis. The quantity of the mRNA in the sample is compared to thequantity of the mRNA in the matched body sample at an earlier time. Areduction of in the quantity of the mRNA after therapy is an index ofefficacy of the therapy. Preferably, the amount of the mRNA is decreasedby at least 10%, 25%, 50%, 75% or 100%.

[0039] According to another aspect of the invention, compounds whichhave anti-chronic mucosal injury or anti-ulcerative colitis activity canbe identified. A colonic cell expressing a gene of the REG family or agene represented by the Hs.111244 polynucleotide can be contacted with atest compound. The test compound can be a pharmacologic agent alreadyknown in the art or can be a compound previously unknown to have anypharmacological activity. The test compound can be naturally occurringor designed in the laboratory. It can be isolated from microorganisms,animals, or plants, and can be produced recombinantly, or synthesized bychemical methods known in the art.

[0040] The cell can be any primary human cell or human cell line whichexpresses a REG family gene, or a gene represented by the Hs.1 11244polynucleotide, as disclosed above. Methods of establishing cultures ofprimary human cells or of culturing cell lines are well known in theart.

[0041] Expression of at least one gene of the REG gene family or thegene represented by the Hs.111244 polynucleotide can be monitored.Expression can be measured in a sample of the same cell populationbefore and after contact with the test compound. Alternatively, controlcell populations can be employed. A test compound which decreasesexpression of at least one member of the REG gene family is identifiedas a potential drug for chronic mucosal injury. A test compound whichdecreases expression of a gene represented by the Hs.111244polynucleotide is identified as a potential drug for decreasingulcerative colitis. Preferably, the test compound decreases the amountof the gene expression product by at least 10%, 25%, 50%, 75% or 100%.

[0042] Sequence Listing

[0043] SEQ ID NO:1 cDNA sequence of pancreatic stone protein (PSP)

[0044] SEQ ID NO:2 cDNA sequence of pancreatitis-associated protein(PAP)

[0045] SEQ ID NO:3 cDNA sequence of human pancreatic beta cell growthfactor (INGAP)

[0046] SEQ ID NO:4 cDNA sequence of regenerating gene homologue (REGH)

[0047] SEQ ID NO:5 cDNA sequence of Hs.11124

EXAMPLE 1

[0048] PSP, PAP and REGH are expressed in colonic mucosa of patientswith inflammatory bowel disease. Parallel methods of measuring geneexpression have been recently developed which allow concurrentmeasurement of the expression of a large number of genes. Light-directedsolid-phase combinatorial chemistry was used to generate oligonucleotideprobe arrays which provide representation of nearly 7000 human cDNA andEST sequences. Each gene is represented by 20 individual 25-meroligonucleotide sequences. mRNA isolated from the mucosa of colonicresection specimens was used to generate hybridization probes for ouranalysis. Details of the GENECHMP technology, probe synthesis,hybridization, and confocal scanning have been previously described. Thefluorescence intensity for different levels of gene expression wasstandardized by spiking known amounts of control genes into the probemixture (FIG. 1). Detection at 1.5 pM is approximately equal to onemessage copy per cell. Tissue samples taken from the area used toisolate RNA were sent for histochemistry to be scored for acute andchronic inflammation, ulceration, dysplasia, eosinophilia, epithelialapoptosis, and metaplastic changes. Expression levels of PSP, PAP, andREGH in 15 clinical specimens are shown in FIG. 2. PSP RNA expressionwas in the top 2% of all arrayed genes in ulcerative colitis. Expressionlevels corresponded closely to histologic measures of disease activity.One non-EBD patient with severe acute inflammation, but no ulceration(rectal prolapse-specimen 11), did not express detectable PSP, PAP,orREGH.

EXAMPLE 2

[0049]Pichia pastoris expression vectors were constructed with PSP, PAP,and REGH. Gene-specific primer pairs were designed to incorporate a 5′Xho I site, and a portion of the a-factor gene leading up to the yeastSte13 cleavage site (5′ end) and a Xba I containing primer that deletedthe stop codon and included a 3′ Myc epitope tag. After Kex2 and Ste13signal cleavage by Pichia, the amino-terminus should be identical to thenative secreted protein. RT-PCR was performed using the TITAN One TubeSystem (Boehringer Mannheim). The PCR products of these reactions, usingRNA from a healthy or UC patient are shown in FIG. 3. These resultsdemonstrate trace amounts of PAP in the “normal” patient examined, butotherwise agree with the results of the GENECHIP hybridization. Bandswere gel purified and cloned into pGEM-T (Promega). The Xba/Xho genefragments were excised and ligated into pPICZ( (Invitrogen). Constructswere bidirectionally sequenced with primers derived from the vector andwere found to match the published sequences. Linearized plasmids weretransformed into Pichia KM71 and recombinant clones identified by Zeocinselection.

EXAMPLE 3

[0050] PSP, PAP, and REGH were expressed in Pichia pastoris. PICZαplaces the inserted gene downstream of a strong methanol-inducible AOX1promoter. Individual clones were grown in 10 ml cultures of BMGY mediaovernight and resuspended into ⅕ volume of BMMY (0.5% methanol) forinduction. Aliquots of media supernatant were taken at various timesafter methanol induction and subjected to 15% SDS-PAGE. PSP or REGHexpression was identified by the new appearance of 18 and 18.5 kDa bands(respectively), peaking 48 hours after induction. These sizes includethe 2.5 kDa C-terminal epitope tag, whose presence was verified byWestern blot (ECL) utilizing a monoclonal anti-Myc antibody(Invitrogen). Tryptic digestion of both PSP and REGH led to a mobilityreduction of about 1 kDa, reflecting the expected size change followingcleavage at the Arg11-Ile12 bond (data not shown). PAP expression wasalso demonstrated.

[0051] Large scale protein purification was performed by directlyscaling up the protocol outlined above. Pichia from 1.5 liter cultureswere resuspended into 300 mls of BMMY induction media and allowed togrow for 48 hours. Culture supernatants were concentrated by ammoniumsulfate precipitation and column purified by Bio-Gel P30. Fractionscontaining PSP or REGH were pooled and concentrated using a Centriprep10 concentrator (Amicon). FIG. 4 shows a Coomassie-stained SDS-PAGE gelof the purified proteins.

1 5 1 777 DNA Homo sapiens 1 ttcttcaaac cctcctcttc cctgtgttct cctacagagattgctgattt ctccttaagc 60 aagagattca ctgccgctaa gcatggctca gaccaactcgttcttcatgc tgatctcctc 120 cctgatgttc ctgtctctga gccaaggcca agaggcccagacagagttgc cccaggcccg 180 gatcagctgc ccagaaggca ccaatgccta tcgctcctactgctactact ttaatgaaga 240 ccgtgagacc tgggttgatg cagatctcta ttgccagaacatgaattcgg gcaacctggt 300 gtctgtgctc acccaggccg agggtgcctt tgtggcctcactgattaagg agagtggcac 360 tgatgacttc aatgtctgga ttggcctcca tgaccccaaaaagaaccgcc gctggcactg 420 gagcagtggg tccctggtct cctacaagtc ctggggcattggagccccaa gcagtgttaa 480 tcctggctac tgtgtgagcc tgacctcaag cacaggattccagaaatgga aggatgtgcc 540 ttgtgaagac aagttctcct ttgtatgcaa gttcaaaaactagaggcagc tggaaaatac 600 atgtctagaa ctgatccagc aattacaacg gagtcaaaaattaaaccgga ccatctctcc 660 aactcaactc aacctggaca ctctcttctc tgctgagtttgccttgttaa tcttcaatag 720 ttttacctac cccagtcttt ggaaccctaa ataataaaaataaacatgtt ttccact 777 2 798 DNA Homo sapiens 2 cgggagagtg actcctgattgcctcctcaa gtcgcagaca ctatgctgcc tcccatggcc 60 ctgcccagtg tatcttggatgctgctttcc tgcctcatgc tgctgtctca ggttcaaggt 120 gaagaacccc agagggaactgccctctgca cggatccgct gtcccaaagg ctccaaggcc 180 tatggctccc actgctatgccttgtttttg tcaccaaaat cctggacaga tgcagatctg 240 gcctgccaga agcggccctctggaaacctg gtgtctgtgc tcagtggggc tgagggatcc 300 ttcgtgtcct ccctggtgaagagcattggt aacagctact catacgtctg gattgggctc 360 catgacccca cacagggcaccgagcccaat ggagaaggtt gggagtggag tagcagtgat 420 gtgatgaatt actttgcatgggagagaaat ccctccacca tctcaagccc cggccactgt 480 gcgagcctgt cgagaagcacagcatttctg aggtggaaag attataactg taatgtgagg 540 ttaccctatg tctgcaaagttcactgacta gtgcaggagg gaagtcagca gcctgtgttt 600 ggtgtgcaac tcatcatgggcatgagacca gtgtgaggac tcaccctgga agagaatatt 660 cgcttaattc ccccaacctgaccacctcat tcttatcttt cttctgtttc ttcctccccg 720 ctagtcattt cagtctcttcattttgtcat acggcctaag gctttaaaga gcaataaaat 780 ttttagtctg caaaaaaa 7983 586 DNA Homo sapiens 3 ttcccatgac cctctgtagg atgtcttgga tgctgctttcctgcctgatg ttcctttctt 60 gggtggaagg tgaagaatct caaaagaaac tgccttcttcacgtataacc tgtcctcaag 120 gctctgtagc ctatgggtcc tattgctatt cactgattttgataccacag acctggtcta 180 atgcagaact atcctgccag atgcatttct caggacacctggcatttctt ctcagtactg 240 gtgaaattac cttcgtgtcc tcccttgtga agaacagtttgacggcctac cagtacatct 300 ggattggact ccatgatccc tcacatggta cactacccaacggaagtgga tggaagtgga 360 gcagttccaa tgtgctgacc ttctataact gggagaggaacccctctatt gctgctgacc 420 gtggttattg tgcagttttg tctcagaaat caggttttcagaagtggaga gattttaatt 480 gtgaaaatga gcttccctat atctgcaaat tcaaggtctagggcagttct aatttcaaca 540 gcttgaaaat attatgaagc tcacatggac aaggaagcaagtatga 586 4 3411 DNA Homo sapiens 4 aggaagggca aagctcaaca tcaacttggacagtttgcca acctgtttgt ggtaagttga 60 tgtcatttgt gaccactcct aatgtgtgccaccaataagc tattcctgat gccagaatct 120 cttactgtca gtgccctctg taggccttctgatccttact ccttgctcca cccattgttt 180 atatcatgta gttctctctc agaccctgatataaagctcc tactctgtct gacctgacaa 240 gccacctcaa gtggacaagg cacttaccaacaggtaaagg ggcattacag gagaagagca 300 tgtctaacgt gggattttct cttttcattttgaggtagat acagggtgat tttctgaata 360 aaagatccca gtagtaatga aacttaagcaagaccaaagc tgatttcggg taatttggcc 420 tctgttatcc ccaaaccaaa agagaaatatctgggagtgt agctatctca gtggaccttt 480 ctgctcacag gaattcagag aggagaggatgttagaaaga taacaggtgc tctgctctct 540 tcttcaaacc ctcttccctg tgttctcctacagagattgc tgatttgctc cttaagcaag 600 agattcactg ccgctaagca tggctcagaccaactcgttc ttcatgctga tctcctccct 660 gatgttcctg tctctgagcc aaggtgagattttcccccac acttcccaca accccaactc 720 tgaattcttc cctccatcct catgtataaggttcacttga aaaaaagcag agtcaacatc 780 agggtttttt tatgttgttc agtgatcattatggctgatt ttatcccatt caaaaacacc 840 ctcaccttca ttcatgggtt tgagacagaatttaatagga ccacttatag gtgaccattg 900 tggttgagtt tatctgattg aatctatatgcgatggcagt ttgggggatg tttttatgta 960 gtcattgcta ggatgagagc taaggcaaacgtgtgcaggg aaaccgagag aaacttgaga 1020 aaggaggaag cctgggtctt taaaggcagaagcctcagcc tcagaattaa aggaaaacga 1080 gaactcattt atttagccta ttcattgtgagctcttgtct tgagcagagg aaactagaga 1140 gaaaagagat aggatgcagg agggcagaagtgagcaatcg ccccagtatt cactgtatcc 1200 atatgttctt ataaggacac caagaagcccctattcacct tccagccttt tccttgccct 1260 gagattcttt cttagttatc tccttttttttttccccagg ccaggagtcc cagacagagc 1320 tgcctaatcc ccgaatcagc tgcccagaaggcaccaatgc ctatcgctcc tactgctact 1380 actttaatga agaccctgag acctgggttgatgcagatgt gagtgaggag agcagcaggg 1440 gaagggaggc ttatgaaggt agaggcagctgctaatttgc agtgtgttct gtggctgcaa 1500 tgagataaga ttgatccctt ccctattccaccactggtcc aaaacttccc aatctacttt 1560 atcccatcat ttgacacatt cccagcacagagatgctggt ggtcagtgac agcatcatca 1620 gggacatttc tgtgctgtcc tttttctgttacatcctctg gaaggtctca gtatatccct 1680 cacaccttcc ttctccactg agtgctccattttcttctcc aacagctcta ttgccagaac 1740 atgaattcag gcaacctggt gtctgtgctcacccaggcgg agggtgcctt cgtggcctca 1800 ctgattaagg agagtagcac tgatgacagcaatgtctgga ttggcctcca tgacccaaaa 1860 aaggtcagtc tgcagccacc tctatctccttataaacatt tttgagaggt aagagggacg 1920 tttaaggtct ggcaccgcaa tcaccaacttttatcttttt gtttgtttaa ataaaagcaa 1980 cctctttata gatcctataa tgtatgagttgtgaagttca gtgtaggtag ttagagacat 2040 gagctgaagg ctgaattttc tgggctctgggaattcatgc acccactcat tgtgtctact 2100 tctagaaatg catctttatg tacaactttttccctatttt gctattgtct gtcttggaag 2160 aggtccctct gtagactata tagaaaatgagtagtggagg agaatctact gctggcattt 2220 gttatacatt ttatacaagt gtataaaactgtacagtata ttatttagtt taacactata 2280 aactaaataa tatatcaaca actactctacagccaatgtt atgctggata tgagagttct 2340 gagattcagg aaaaaaatca gaaaccactctctgtaatgg gcttttatgg gtctctgtat 2400 caaattctga acacttatta tttgctagaagaggaggagg aattcggaca ttctagagaa 2460 ggagaagctt agagcaaaag cagaggaaatgatatgatat tcatggtgac aacaatgttt 2520 attctttctg ctataacttg gcctgtttctgagtgtgcac acaggcctgg ttattctatt 2580 gatttttgag tgaccatggc ccctgttctggcccttctcc atctagaacc gccgctggca 2640 ctggagtagt gggtccctgg tctcctacaagtcctgggac actggatccc cgagcagtgc 2700 taatgctggc tactgtgcaa gcctgacttcatgctcaggt gagaggcaga caatctatcc 2760 acctgttgcc atttccttcc cacttatctctggggatgaa catggggact gggatagagg 2820 aaaggtaagc tccttatctg gaaaataaagaagtatttcc tctagttttt tgttctgagt 2880 cctaggttga ggaggggcta cactccttctgatcctctat gtctgacact tctcattgta 2940 ctataggatt caagaaatgg aaggatgaatcttgtgagaa gaagttctcc tttgtttgca 3000 agttcaaaaa ctagaggaag ctgaaaaatggatgtctaga actggtcctg caattactat 3060 gaagtcaaaa attaaactag actatgtctccaactcagtt cagaccatct cctccctaat 3120 gagtttgcat cgctgatctt cagtaccttcacctgtctca gtctctagag ccctgaaaaa 3180 taaaaacaaa cttattttta tccagtgttctgtcttctgc atttgctctt tctacagccc 3240 atgcttgggt ggttggggtg ggaatgattgtcacactcca gagcttgcca tggcccatcc 3300 acttgttaaa accccactca cattttatgtatgtcaggct tatgaacatg tggtggcctt 3360 gtttatgaca agataaaaag attaagatttcatccacaac acatgttagc a 3411 5 1734 DNA Homo sapiens 5 gcgagcgtggacctgggacg ggtctgggcg gctctcggtg gttggcacgg gttcgcacac 60 ccattcaagcggcaggacgc acttgtctta gcagttctcg ctgaccgcgc tagctgcggc 120 ttctacgctccggcactctg agttcatcag caaacgccct ggcgtctgtc ctcaccatgc 180 ctagcctttgggaccgcttc tcgtcgtcgt ccacctcctc ttcgccctcg tccttgcccc 240 gaactcccaccccagatcgg ccgccgcgct cagcctgggg gtcggcgacc cgggaggagg 300 ggtttgaccgctccacgagc ctggagagct cggactgcga gtccctggac agcagcaaca 360 gtggcttcgggccggaggaa gacacggctt acctggatgg ggtgtcgttg cccgacttcg 420 agctgctcagtgaccctgag gatgaacact tgtgtgccaa cctgatgcag ctgctgcagg 480 agagcctggcccaggcgcgg ctgggctctc gacgccctgc gcgcctgctg atgcctagcc 540 agttggtaagccaggtgggc aaagaactac tgcgcctggc ctacagcgag ccgtgcggcc 600 tgcggggggcgctgctggac gtctgcgtgg agcagggcaa gagctgccac agcgtgggcc 660 agctggcactcgaccccagc ctggtgccca ccttccagct gaccctcgtg ctgcgcctgg 720 actcacgactctggcccaag atccaggggc tgtttagctc cgccaactct cccttcctcc 780 ctggcttcagccagtccctg acgctgagca ctggcttccg agtcatcaag aagaagctgt 840 acagctcggaacagctgctc attgaggagt gttgaacttc aacctgaggg ggccgacagt 900 gccctccaagacagagacga ctgaactttt ggggtggaga ctagaggcag gagctgaggg 960 actgattcctgtggttggaa aactgaggca gccacctaag gtggaggtgg gggaatagtg 1020 tttcccaggaagctcattga gttgtgtgcg ggtggctgtg cattggggac acatacccct 1080 cagtactgtagcatgaaaca aaggcttagg ggccaacaag gcttccagct ggatgtgtgt 1140 gtagcatgtaccttattatt tttgttactg acagttaaca gtggtgtgac atccagagag 1200 cagctgggctgctcccgccc cagcccggcc cagggtgaag gaagaggcac gtgctcctca 1260 gagcagccggagggaggggg gaggtcggag gtcgtggagg tggtttgtgt atcttactgg 1320 tctgaagggaccaagtgtgt ttgttgtttg ttttgtatct tgtttttctg atcggagcat 1380 cactactgacctgttgtagg cagctatctt acagacgcat gaatgtaaga gtaggaaggg 1440 gtgggtgtcagggatcactt gggatctttg acacttgaaa aattacacct ggcagctgcg 1500 tttaagccttcccccatcgt gtactgcaga gttgagctgg caggggaggg gctgagaggg 1560 tgggggctggaacccctccc cgggaggagt gccatctggg tcttccatct agaactgttt 1620 acatgaagataagatactca ctgttcatga atacacttga tgttcaagta ttaagaccta 1680 tgcaatattttttacttttc taataaacat gtttgttaaa acaaaaaaaa aaaa 1734

1. A method of diagnosing chronic inflammatory bowel disease comprising:detecting at least one gene expression product of the regenerating (EG)gene family in a body sample of a first human, wherein the first humanis suspected of having chronic inflammatory bowel disease; identifyingthe first human as having chronic inflammatory bowel disease if the geneexpression product is detected.
 2. The method of claim 1 wherein anamount of the gene expression product detected in the body sample of thefirst human is compared with an amount of the gene expression productdetected in a body sample of a second human, wherein the second human ishealthy, wherein more of the gene expression product detected in thebody sample of the first human than in the body sample of the secondhealthy human, confirms chronic mucosal injury in the first human. 3.The method of claim 1 wherein the gene expression product of the REGgene family is selected from the group consisting of gene expressionproducts of pancreatic stone protein (PSP), pancreatitis associatedprotein (PAP), human pancreatic beta cell growth factor (INGAP), andregenerating gene homologue (REGH) genes.
 4. The method of claim 1wherein the chronic mucosal injury is selected from the group ofdiseases consisting of ulcerative colitis and Crohn's disease.
 5. Themethod of claim 1 wherein the body sample is blood.
 6. The method ofclaim 1 wherein the body sample is plasma.
 7. The method of claim 1wherein the body sample is serum.
 8. The method of claim 1 wherein thebody sample is small intestine or colon tissue.
 9. The method of claim 1wherein the gene expression product is a polypeptide.
 10. The method ofclaim 9 wherein an antibody is used to detect the polypeptide.
 11. Themethod of claim 10 wherein an assay selected from the group consistingof Western blot assay, immunoprecipitation assay, enzyme linedimmunoabsorbant assay, quantitative antigen capture-based immunoassay,and radioimmunoassay is used to detect the polypeptide.
 12. The methodof claim 1 wherein the gene expression product is mRNA.
 13. The methodof claim 12 wherein an assay selected from the group consisting ofNorthern blot assay, DNA array, and ribonuclease protection assay, isused to detect the mRNA.
 14. A method to aid in the differentiation ofchronic mucosal injury from common acute inflammatory colon disorder andcommon non-inflammatory benign colon disorder in a human with symptomsof bowel disease comprising: comparing (a) the amount of at least onegene expression product of the REG gene family in a body sample of afirst human who is suspected of having bowel disease, with (b) theamount of the gene expression product in a body sample of a second humanwho is healthy; identifying the first human as having chronic mucosalinjury if the body sample of the first human contains more of the geneexpression product than the body sample of the second human.
 15. Themethod of claim 14 wherein the gene expression product of the REG genefamily is selected from the group consisting of gene expression productsof pancreatic stone protein (PSP), pancreatitis-associated protein(PAP), human pancreatic beta cell growth factor (7NGAP), andregenerating gene homologue UGH) genes.
 16. The method of claim 14wherein the body sample is blood.
 17. The method of claim 14 wherein thebody sample is plasma.
 18. The method of claim 14 wherein the bodysample is serum.
 19. The method of claim 14 wherein the body sample issmall intestine or colon tissue.
 20. The method of claim 14 wherein thegene expression product is a polypeptide.
 21. The method of claim 20wherein an antibody is used to quantitate the polypeptide.
 22. Themethod of claim 21 wherein an assay selected from the group consistingof Western blot assay, immunoprecipitation assay, enzyme linkedimmunoabsorbant assay, quantitative antigen capture-based immunoassay,and radioimmunoassay is used to quantitate the polypeptide.
 23. Themethod of claim 14 wherein the gene expression product is mRNA.
 24. Themethod of claim 23 wherein an assay selected from the group consistingof Northern blot assay, DNA array, and ribonuclease protection assay, isused to detect the mRNA.
 25. A method to determine degree of injury tosmall intestine or colon tissue of a human with chronic mucosal injurycomprising the steps of: determining a quantity of a gene expressionproduct of the REG gene family in a body sample of a human havingchronic mucosal injury, correlating the quantity of the gene expressionproduct with the degree of injury to the small intestine or colon. 26.The method of claim 25 wherein the gene expression product of the REGgene family is selected from the group consisting of gene expressionproducts of pancreatic stone protein (PSP), pancreatitis-associated in(PAP), human pancreatic beta cell growth factor (INGAP), andregenerating gene homologue (REGH) genes.
 27. The method of claim 25wherein the chronic mucosal injury is selected from the group ofdiseases consisting of ulcerative colitis and Crohn's disease.
 28. Themethod of claim 25 wherein the body sample is blood.
 29. The method ofclaim 25 wherein the body sample is plasma.
 30. The method of claim 25wherein the body sample is serum.
 31. The method of claim 25 wherein thebody sample is small intestine or colon tissue.
 32. The method of claim25 wherein the gene expression product is a polypeptide.
 33. The methodof claim 32 wherein an antibody is used to quantitate the polypeptide.34. The method of claim 33 wherein an assay selected from the groupconsisting of Western blot assay, immunoprecipitation assay, enzymelinked immunoabsorbant assay, quantitative antigen capture-basedimmunoassay, and radioimmunoassay is used to quantitate the polypeptide.35. The method of claim 25 wherein the gene expression product is mRNA.36. The method of claim 35 wherein an assay selected from the groupconsisting of Northern blot assay, DNA array, and ribonucleaseprotection assay is used to detect the
 37. A method of monitoring theefficacy of therapy for chronic mucosal injury in a human body samplecomprising the steps of: quantitating at least one gene expressionproduct of the REG gene family in a body sample of a human who has beensubjected to therapy for chronic mucosal injury; comparing the quantityof expression product in said sample to the quantity of said geneexpression product in a matched body sample of the human at an earliertime, wherein a reduction in the quantity of said gene expressionproduct after therapy is an index of efficacy of the therapy.
 38. Themethod of claim 37 wherein the gene expression product of the REG genefamily is selected from the group consisting of gene expression productsof pancreatic stone protein (PSP), pancreatitis-sociatedprotein (PAP),human pancreatic beta cell growth factor (INGAP), and regenerating genehomologue (REGH) genes.
 39. The method of claim 37 wherein the chronicmucosal injury is selected from the group of diseases consisting ofulcerative colitis and Crohn's disease.
 40. The method of claim 37wherein the body sample is blood.
 41. The method of claim 37 wherein thebody sample is plasma.
 42. The method of claim 37 wherein the bodysample is serum.
 43. The method of claim 37 wherein the body sample issmall intestine or colon tissue.
 44. The method of claim 37 wherein thegene expression product is a polypeptide.
 45. The method of claim 44wherein an antibody is used to quantitate the polypeptide.
 46. Themethod of claim 45 wherein an assay selected from the group consistingof Western blot assay, immunoprecipitation assay, enzyme linkedimmunoabsorbant assay, quantitative antigen capture-based immunoassay,and radioimmunoassay is used to quantitate the polypeptide.
 47. Themethod of claim 37 wherein the gene expression product is mRNA.
 48. Themethod of claim 47 wherein an assay selected from the group consistingof Northern blot assay, DNA array, and ribonuclease protection assay isused to detect the mRNA.
 49. A method of screening compounds foranti-chronic mucosal injury activity comprising: contacting a coloniccell expressing a gene which is a member of the REG gene family with atest compound and; quantitating expression of the REG gene, wherein atest compound which decreases expression of the gene is identified as apotential compound for treating chronic mucosal injury.
 50. The methodof claim 49 wherein the gene is selected from the group consisting ofpancreatic stone protein (PSP), pancreatitis-associated protein (PAP),human pancreatic beta cell growth factor (INGAP), and regenerating genehomologue (REGH) genes.
 51. A method of diagnosing ulcerative colitiscomprising: detecting an mRNA which is expressed by a gene representedby a Hs.111244 polynucleotide in a body sample of a first human who issuspected of having ulcerative colitis; identifying the human as havingulcerative colitis if said mRNA is detected.
 52. The method of claim 51wherein an amount of the mRNA detected in the body sample of the firsthuman is compared with an amount of the mRNA in a body sample of asecond human who is healthy, identifying the first human as havingulcerative colitis if the body sample of the first human contains moreof the mRNA than the body sample of the second human.
 53. The method ofclaim 51 wherein the body sample is blood.
 54. The method of claim 51wherein the body sample is plasma.
 55. The method of claim 51 whereinthe body sample is serum.
 56. The method of claim 51 wherein the bodysample is small intestine or colon tissue.
 57. The method of claim 51wherein an assay selected from the group consisting of Northern blotassay, DNA array, and ribonuclease protection assay is used to detectthe mRNA.
 58. A method to aid in the differentiation of ulcerativecolitis from common acute inflammatory colon disorder, commonnon-inflammatory benign colon disorder, and Crohn's disease in a humanwith symptoms of bowel disease comprising: comparing the amount of mRNAwhich is expressed by a gene represented by a Hs.111244 polynucleotidein a body sample of a first human suspected of having bowel disease withthe amount of the mRNA in a comparable body sample of a second human whois healthy, wherein a body sample of the first human which contains moreof the mRNA than the body sample of the second human identifies thefirst human as having ulcerative colitis.
 59. The method of claim 58wherein the body sample is blood.
 60. The method of claim 58 wherein thebody sample is plasma.
 61. The method of claim 58 wherein the bodysample is serum.
 62. The method of claim 58 wherein the body sample issmall intestine or colon tissue.
 63. The method of claim 58 wherein anassay selected from the group consisting of Northern blot assay, DNAarray, and ribonuclease protection assay is used to detect the mRNA. 64.A method to determine degree of injury to small intestine or colontissue of a human with ulcerative colitis comprising the steps of:determining a quantity of an mRNA which is expressed by a generepresented by a Hs.111244 polynucleotide in a body sample of a firsthuman having ulcerative colitis; correlating the quantity of the mRNAwith the degree of injury to the small intestine or colon.
 65. Themethod of claim 64 wherein the body sample is blood.
 66. The method ofclaim 64 wherein the body sample is plasma.
 67. The method of claim 64wherein the body sample is serum.
 68. The method of claim 64 wherein thebody sample is small intestine or colon tissue.
 69. The method of claim64 wherein an assay selected from the group consisting of Northern blotassay, DNA array, and ribonuclease protection assay is used to detectthe mRNA.
 70. A method of monitoring the efficacy of therapy forulcerative colitis in a human body sample comprising the steps of:quantitating an mRNA which is expressed by gene represented by aHs.111244 polynucleotide in a body sample of a human who has beensubjected to therapy for ulcerative colitis; comparing the quantity ofthe mRNA in said sample to the quantity of said mRNA in a matched bodysample of the human at an earlier time, wherein a reduction in thequantity of said mRNA after therapy is an index of efficacy of thetherapy.
 71. The method of claim 70 wherein the body sample is blood.72. The method of claim 70 wherein the body sample is plasma.
 73. Themethod of claim 70 wherein the body sample is serum.
 74. The method ofclaim 70 wherein the body sample is small intestine or colon tissue. 75.The method of claim 70 wherein an assay selected from the groupconsisting of Northern blot assay, DNA array, and ribonucleaseprotection assay is used to detect the mRNA.
 76. A method of screeningcompounds for anti-ulcerative colitis activity comprising: contacting acolonic cell expressing an mRNA which is expressed by a gene representedby a Hs.111244 polynucleotide with a test compound and; quantitatingexpression of the mRNA by the cell, wherein a test compound whichdecreases expression of the mRNA is identified as a potential compoundfor treating ulcerative colitis.